The overall purpose of this proposal is to examine the balance between nitric oxide (NO) production and superoxide-mediated NG destruction in the normal and hypoxic/hypertensive pulmonary circulation, in an effort to establish genes of interest for gene transfer in pulmonary hypertension (PHTN). NO is an important inhibitory modulator of pulmonary artery tone and medial smooth muscle cell proliferation. During the development of PHTN, endogenous NO activity is insufficient to completely oppose vasoconstriction and vascular wall remodeling. This relative insufficiency of NO activity occurs despite increased expression of endothelial constitutive nitric oxide synthase (eNOS), suggesting that counter regulatory factors may inhibit endogenous eNOS or accelerate destruction of NO. An important mechanism through which NO may be destroyed is reaction with superoxide anion. This reaction can be prevented by superoxide dismutase (SOD). We hypothesize that during the development of PHTN, insufficient NOS and SOD activity may independently or in combination lead to submaximal bioavailable NO, resulting in pulmonary vasoconstriction and vascular wall remodeling. We therefore propose to test the hypothesis that reduction in pulmonary NOS or SOD activity will enhance pulmonary vasoconstriction using mice with targeted deletion of either eNOS, inducible (i)NOS, extracellular (EC-SOD, or Cu,Zn-SOD. We will also test the converse hypothesis, that pulmonary NO production can be augmented, and pulmonary vasoconstriction attenuated, in vivo, by transient overexpression of NOS (either eNOS or iNOS), and SOD (either EC-SOD or Cu,Zn-SOD). Expression will be targeted to pulmonary endothelium by injection of lung-avid cationic lipid/DNA complexes. Using the combined approaches of knockout and transient overexpression of the genes of interest, we will more definitively establish the roles of NOS and SOD in modulating the development of PHTN than has previously been possible using pharmacological approaches. While our focus includes a detailed physiological evaluation of the roles of NOS and SOD in modulating pulmonary vascular tone, our overall goal is development of novel therapies for humans with PHTN. Thus, testing if NOS and/or SOD overexpression attenuates pulmonary vasoconstriction will serve both to identify potential genes of interest for "gene therapy" for PHTN, as well as providing proof of principle that pulmonary vascular gene transfer could merit future study as a novel therapeutic approach in PHTN.